CRYSTAL STRUCTURES OF SOME PROLYL-OLIGOPEPTIDASE - INHIBITOR COMPEXES

V. Harmat1, Zs. Böcskei1, V. Fülöp2, K. Kánai3, G. Náray-Szabó1, I. Hermecz3, L. Polgár4, P. Arányi3

1Department of Theoretical Chemistry, Etvs University, Budapest
2Laboratoy of Molecular Biophysics, University of Oxford
3CHINOIN Pharmaceuticals
4Institute of Enzymology, Biological Research Center of the HAS, Budapest

Prolyl-oligopeptidase (PEP) is an intracellular serine peptidase which cleaves peptide bonds at the carboxyl side of prolyl residues [1]. It acts only on smaller polypeptides or oligopeptides. The structure of PEP [2] shows a C terminal peptidase domain evolved on an a /b -hydrolase fold while the N terminal b -propeller domain regulates the access to the active site. The active site is in the middle of the molecule in a huge cavity.

PEP is assumed to be involved in the control of the blood pressure and in the process of learning and memory by playing role in the metabolism of peptide hormones and neuropeptides [3 - 5]. Therefore its inhibitors are potential drugs in the treatment of amnesia.

Molecules containing prolyl-pyrrolidine groups have inhibitory effect since their peptide bond connecting the two five - membered rings cannot dock on the active site due to steric clashes between the C terminal ring and nearby residues. This forces the C-terminal ring to occupy the main substrate specificity pocket, while the side chain of the N-terminal Pro points toward the large central cavity. The third fragments of the inhibitors connected to the N-terminal Pro are bound in a hydrophobic pocket. The strength of binding can be further increased by introduction of formyl or keto groups at the C-terminal side resulting in covalently binding inhibitors.

We investigated the structure of PEP-inhibitor complexes to map the enzyme - inhibitor interactions responsible for drug binding. We obtained crystals of several PEP - inhibitor complexes (including both covalently and noncovalently binding molecules) by co-crystallization using the microseeding techique [6]. The analysis of the complex structures helped us to interpret the results of the kinetic studies and to suggest modifications of the inhibitor structures.

  1. Polgár, L. (1992) FEBS Letters 311, 281-284
  2. Fülöp,V., Böcskei, Zs. & Polgár, L. (in preparation)
  3. Polgár, L. (1994) Methods in Enzymol 244, 188-200
  4. Wilk, S. (1983) Life Sci. 33, 2149-2157
  5. Yosimoto,T., Kado,K., Matsubara,F., Koriyama,N., KanettoH. & Tsuru,D. (1987) J. Phamacobio-Dyn. 10, 730-735
  6. Böcskei,Zs., Fuxreiter,M., Náray-Szabó,G., Szabó,E. & Polgár, L., Acta Cryst. D. (in press)